Monocyclic functional dyes for contrast enhancement in optical imaging

ABSTRACT

This invention provides functional dyes of the general formula: ##STR1## wherein R 1 , R 2 , and R 5  may be the same or different and are selected from the group consisting of hydrogen, C 1  -C 10  alkyl, C 1  -C 10  alkoxyl, hydroxyl, C 1  -C 10  hydroxyalkyl, C 1  -C 10  alkoxyalkyl, C 1  -C 10  aryl, carboxyl, C 1  -C 10  carboxylalkyl, halogen, nitro, C 1  -C 10  alkoxycarbonyl, mercapto, C 1  -C 10  mercaptoalkyl, C 1  -C 10  alkylthio, sulfonate, and --(CH 2 ) m  --N(R 6 )(R 7 ) wherein R 6  and R 7  are independently hydrogen or C 1  -C 10  alkyl, C 1  -C 10  acyl, and R 6  and R 7  are capable of forming 5, 6, or 7 membered rings which may optionally be substituted with --O--, --NR 8 , or --S--; R 3  and R 4  may be the same or different and are selected from the group consisting of C 1  -C 10  alkyl, C 1  -C 10  hydroxyalkyl, C 1  -C 10  alkoxyalkyl, C 1  -C 10  aryl, C 1  -C 10  carboxylalkyl, C 1  -C 10  alkyl sulfonate, mercapto alkyl and --(CH 2 ) m  N(R 6 )(R 7 ); X is --O--, --NR 8 , or --S--; W 1  and W 2  may be the same or different and are selected from the group consisting of --S--, --O--, --NR 8  and C(R 9 )(R 10 ); p is about 1-10; and m is about 0-10; R 8  is hydrogen, C 1  -C 10  alkyl, C 1  -C 10  alkoxy, C 1  -C 10  mercaptoalkyl, hydroxyl, C 1  -C 10  hydroxyalkyl, C 1  -C 10  alkoxyalkyl, C 1  -C 10  aryl, C 1  -C 10  carboxylalkyl, C 1  -C 10  alkoxycarbonyl, C 1  -C 10  alkylthio, and --(CH 2 ) m  --N(R 6 )(R 7 ); and R 9  and R 10  are independently hydrogen, C 1  -C 10  alkyl, C 1  -C 10  alkoxyl, C 1  -C 10  hydroxyalkyl, C 1  -C 10  alkoxyalkyl, C 1  -C 10  carboxyalkyl, C 1  -C 10  alkoxycarbonyl, and --(CH 2 ) m  --N(R 6 )(R 7 ).

FIELD OF THE INVENTION

The invention is in the field of tomographic imaging. Particularly, theinvention is in the field of optical imaging. Most particularly, theinvention provides dyes for use in the field of optical imaging.

BACKGROUND OF THE INVENTION

Optical imaging with dyes permit visualization of biological activities(Blasdel, G. G.; Salama, G. Nature 1986, 321, 579, Grinvald, A.;Frostig, E. L.; Hildesheim, R. Physiological Reviews 1988, 68, 1285,Kauer, J. S. Nature 1988, 331, 166, Lieke, E. E.; Frostig, R. D.;Arieli, A.; Ts'o, D. Y.; Hildesheim, R. and Grinvald, A. Annu. Rev.Physiol. 1989, 51, 543 and reference therein). Dyes that are sensitiveto physicochemical environments (such as pressure, cell membranepotential, ion concentration, acidity, partial pressure of oxygen andetc.), are subject to changes in absorption or emission of light. Theresulting changes act as optical probes to transform biologicalactivities into optical signals that can be converted into opticalimages.

Cyanine dyes with intense absorption and emission in the near-IR region(600-1300 nm) are particularly useful because biological tissues areoptically transparent in this region. Indocyanine Green (ICG) (I) forexample, with absorption maxima at around 810 nm (the isosbestic pointof the hemoglobin/deoxyhemoglobin), has been used for monitoring cardiacoutput, hepatic function, and liver blood flow. After intravenousinjection, ICG is rapidly bound by plasma proteins and remains in theblood through one circulation of heart and lungs. ICG is then taken upby hepatic parenchymal cells and secreted entirely into the bile (Osol,A.; Pratt, R. The United States Dispensatory Philadelphia, Toronto: I.B. Lippincott Company, 1973, 615).

Despite ICG's promising application, aqueous solutions of indocyaninegreen rapidly decomposes when irradiated with incandescent light. Also,ICG itself does not localize in any particular tissue. ##STR2##

Targeting groups can be introduced to cyanine and indocyanine dyes ifessential linkers are present at a convenient site that will notinterfere with the optical activity. Conventionally, these spacers havebeen attached at the nitrogen atom in the heterocyclic moiety.(Mujumdar, R. B.; Ernst, L. A.; Mujumdar, S. R.; Lewis, C. J.; Waggoner,A. S. Bioconjugate Chem. 1993, 4, 105). To effectively label targetinggroups, a single spacer between the dye and the targeting group ispreferred, and typically involves a multi-step synthesis (Mujumdar, R.B.; Ernst, L. A.; Mujumdar, S. R.; Lewis, C. J.; Waggoner, A. S.Bioconjugate Chem. 1993, 4, 105). Additional problems also arise fromthe photo-instability of the long olefin chain (Matsuoka, M. In InfraredAbsorbing Dyes; Plenum: New York, 1990; Chapter 3). Cyanine dyes withshorter olefin chains are relatively stable, but their absorption andemission do not fall within the optical window, (600-1300 nm) necessaryfor optical imaging.

There is a need to design stable dyes that possess desirablephotophysical properties, stability, and targeting ability. The presentinvention overcomes the technical problems mentioned previously byincorporating a bifunctional spacer, and imposing rigidity on thepolyene portion of the cyanine and indocyanine dyes.

SUMMARY OF THE INVENTION

This invention provides functional dyes of the general formula: ##STR3##wherein R¹, R², and R⁵ may be the same or different and are selectedfrom the group consisting of hydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀ alkoxyl,hydroxyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ alkoxyalkyl, C₁ -C₁₀ aryl,carboxyl, C₁ -C₁₀ carboxylalkyl, halogen, nitro, C₁ -C₁₀ alkoxycarbonyl,mercapto, C₁ -C₁₀ mercaptoalkyl, C₁ -C₁₀ alkylthio, sulfonate, and--(CH₂)_(m) --N(R⁶)(R⁷) wherein R⁶ and R⁷ are independently hydrogen orC₁ -C₁₀ alkyl, C₁ -C₁₀ acyl, and R⁶ and R⁷ are capable of forming 5, 6,or 7 membered rings which may optionally be substituted with --O--,--NR⁸, or --S--; R³ and R⁴ may be the same or different and are selectedfrom the group consisting of C₁ -C₁₀ alkyl, C₁ -C₁₀ hydroxyalkyl, C₁-C₁₀ alkoxyalkyl, C₁ -C₁₀ aryl, C₁ -C₁₀ carboxylalkyl, C₁ -C₁₀ alkylsulfonate, mercapto alkyl and --(CH₂)_(m) N(R⁶)(R⁷); X is --O--, --NR⁸,or --S--; W¹ and W² may be the same or different and are selected fromthe group consisting of --S--, --O--, --NR⁸ and C(R⁹)(R¹⁰); p is about1-10; and m is about 0-10; R⁸ is hydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀alkoxy, C₁ -C₁₀ mercaptoalkyl, hydroxyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀alkoxyalkyl, C₁ -C₁₀ aryl, C₁ -C₁₀ carboxylalkyl, C₁ -C₁₀alkoxycarbonyl, C₁ -C₁₀ alkylthio, and --(CH₂)_(m) --N(R⁶)(R⁷); and R⁹and R¹⁰ are independently hydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀ alkoxyl, ₁-C₁₀ hydroxyalkyl, C₁ -C₁₀ alkoxyalkyl, C₁ -C₁₀ carboxyalkyl, C₁ -C₁₀alkoxycarbonyl, and (CH₂)_(m) --N(R⁶)(R⁷).

Also provided are methods of using dyes of the invention comprisingadministering a diagnostically effective amount of the dye to a patientand visualizing the dye.

DETAILED DESCRIPTION OF INVENTION

Cyanine dyes are symmetric molecules with two heterocyclic base groupslinked by a conjugated carbon chain. These dyes absorb intensely fromthe visible to near-infrared (NIR) region, depending strongly on thelength of the carbon chain. Bathochromic shift is often found when nincreases (Matsuoka, M. In Infrared Absorbing Dyes; Plenum: New York,1990; Chapter 2 & 3). Other structural variations that causebathochromic shift can also be introduced into these dyes. Typically,electron donating or electron withdrawing functional groups such asalkyl, alkoxyl, alkylthio, halogen, cyano, alkoxycarbonyl, and nitro canbe substituted at the appropriate electron-rich or electron deficientcenters as the polyene portion of the molecule.

These dyes are generally prepared from condensation reaction betweenquaternary salts of heterocyclic base and bis-aldehyde or Schiff base inacetic acid/acetic anhydride (Keyes, G. H. U.S. Pat. No. 2,251,286,1941; Heseltine, D. W.; Brooker, L. G. S. U.S. Pat. No. 2,895,955, 1959;Reynolds, G. A.; K. H. J. Org. Chem. 1977, 42, 885). Despite the successwith this procedure, purification of products is difficult. Asymmetricdyes with linkage groups can be prepared in a similar manner withadditional synthetic steps (Mujumdar, R. B.; Ernst, L. A.; Mujumdar, S.R.; Lewis, C. J.; Waggoner, A. S. Bioconjugate Chem. 1993, 4, 105).

Recently, Narayanan and Patonay have demonstrated the synthesis of newcyanine dyes using 2-chloro-1-formyl-3-(hydroxymethylene) cyclohex-1-eneas a template (Narayanan, N.; Patonay, G. J. Org. Chem. 1995, 60, 2391).Several symmetric dyes were prepared in high yields. Like croconiumdyes, the central part of these dyes contain a cyclic ring with one C═Cbond and a chloride atom. Their results show that introduction of acroconic moiety into the conjugating bridge significantly shift theabsorption to longer wavelengths.

The present invention describes the use of ring template strategy tosynthesize stable cyanine dyes with desirable photophysical andtargeting properties. Simple condensation between a quaternary salt of aheterocyclic base and the template ring will give desirable dye productsthat are conformationally rigid. The extended conjugated π systemthrough this centrally located ring(s) will give intensely coloredmaterials. With appropriate coupling groups such as acid halides, activeesters, alcohols, aldehydes, amines, aryl halides, carboxylic acids,n-carboxyanhydrides, disulfides, hydrazides, iodacetamides,isothiocyanates, imadates, maleimides, nitrenes, sulfonyl chlorides andso forth, the dye moiety can effectively label biological materials. Thering system may have at least one C═C bond. The ring system may alsocontain Group III, IV, V or VI elements in order to further shift theabsorption maxima to longer wavelength. The ring system can also be afused ring structure with five, six or seven membered rings. The ringstructure can also be substituted with acid halides, activer esters,alcohols, aldehydes, amines, aryl halides, carboxylic acids,n-carboxyanhydrides, disulfides, hydrazides, iodoacetamides,isothiocyanates, imadates, maleimides, nitrenes, sulfonyl chloride andso forth that are essential for conjugation with targeting groups(biomolecules).

Biomolecules for use with the dyes refer to all natural and syntheticmolecules that play a role in biological systems. Biomolecules includehormones, amino acids, peptides, peptidomimetics, glycomimetics,vitamins, carbohydrates, proteins, deoxyribonucleic acid (DNA),ribonucleic acid (RNA), lipids, albumins, polyclonal antibodies,receptor molecules, receptor binding molecules, monoclonal antibodiesand aptamers. Specific examples of biomolecules include insulins,prostaglandins, growth factors, liposomes and nucleic acid probes.Examples of synthetic polymers include polylysine, aborols, dendrimers,and cyclodextrins. The advantages of using biomolecules include enhancedtissue targeting through specificity and delivery. Coupling of the dyesto biomolecules can be accomplished by several known methods (e.g.,Krejcarek and Tucker Biochem. Biophys. Res. Comm, 30, 581 (1977);Hnatowich, et at. Science, 220, 613 (1983). Typically, a nucleophilicgroup is reacted with an electrophilic group to form a covalent bondbetween the biomolecule and the dye. Examples of nucleophilic groupsinclude amines, anilines, alcohols, phenols, thiols and hydrazines.Electrophilic group examples include halides, disulfides, epoxides,maleimides, acid chlorides, anhydrides, mixed anhydrides, activatedesters, imidates, isocyanates and isothiocyanates.

Examples of suitable alkyl groups for use with the invention includemethyl, ethyl, propyl, isopropyl, butyl, cyclohexyl, heptyl and octyl.Suitable alkoxyl groups include methoxyl, ethoxyl, propoxyl, butoxyl,pentoxyl, hexoxyl, heptoxyl and octoxyl. Hydroxyalkyl groups suitablefor use with the invention include both mono and poly hydroxyalkyls suchas hydroxyethyl, 2-hydroxypropyl, 2,3-dihydroxypropyl,2,3,4-trihydroxybutyl, tris (hydroxymethyl) methyl and2-hydroxy-1-hydroxymethyl-ethyl. Suitable alkoxyalkyl groups includemethoxymethyl, 2,3-dimethoxypropyl, tris (methoxymethyl) methyl, and2-methoxy-1-methoxymethyl-ethyl. Amino groups suitable for use with theinvention include aminoalkyl such as amino methyl, amino ethyl, aminopropyl, hydroxyamino such as 1-amino-2,3 propandiol, 1-amino-2-ethanol,and 1-amino-3-propanol and amino acids such as alanine, aspartic acid,glycine and lysine. Carboxyalkyls include acetate, hexanoate,propionate, and butyrate. Carbohydrates, monosaccharides, andpolysaccharides such as glucose, maltose, lactose and amylose. Arylgroups include phenyl and naphthyl. Alkoxycarbonyl include methyl ester,ethyl ester, propyl ester and butyl ethyl ester. Halogen groups includechlorine, fluorine, bromine and iodine. Alkylamido groups include groupssuch as methyl amido, ethyl amido, propyl amido and, butyl amide.Alkylthio groups include methyl thio, ethyl thio, propyl thio, and butylthio. Aminoalkyls include NR⁶ R⁷ where R⁶ and R⁷ can be hydrogen or C₁-C₁₀ alkyl and R⁶ and R⁷ are capable of forming 5, 6, or 7 memberedrings which can be further substituted by a heteroatom such as O, --NR⁸or S, wherein R⁸ is hydrogen, alkyl, alkoxyl, hydroxyl, hydroxyalkyl,aminoalkyl, alkoxyalkyl, alkylamido, aryl, carboxyl, carboxyalkyl,halogen, nitro, alkoxycarbonyl, mercapto, alkylthio and sulfonate.

The compositions of the invention can be formulated into diagnosticcompositions for enteral or parenteral administration. Thesecompositions contain an effective amount of the dye along withconventional pharmaceutical carriers and excipients appropriate for thetype of administration contemplated. For example, parenteralformulations advantageously contain a sterile aqueous solution orsuspension of dye according to this invention. Parenteral compositionsmay be injected directly or mixed with a large volume parenteralcomposition for systemic administration. Such solutions also may containpharmaceutically acceptable buffers and, optionally, electrolytes suchas sodium chloride.

Formulations for enteral administration may vary widely, as is wellknown in the art. In general, such formulations are liquids whichinclude an effective amount of the dye in aqueous solution orsuspension. Such enteral compositions may optionally include buffers,surfactants, thixotropic agents, and the like. Compositions for oraladministration may also contain flavoring agents and other ingredientsfor enhancing their organoleptic qualities.

The diagnostic compositions are administered in doses effective toachieve the desired enhancement. Such doses may vary widely, dependingupon the particular dye employed, the organs or tissues which are thesubject of the imaging procedure, the imaging procedure, the imagingequipment being used, and the like.

The diagnostic compositions of the invention are used in theconventional manner. The compositions may be administered to a patient,typically a warm-blooded animal, either systemically or locally to theorgan or tissue to be imaged, and the patient then subjected to theimaging procedure.

The following examples illustrate the specific embodiments of theinvention described in this document. As would be apparent to skilledartisans, various changes and modifications are possible and arecontemplated within the scope of the invention described.

EXAMPLES

Example I

Synthesis of Dimethylbenzothiazolium monocarbothiophene iodide

A mixture of 0.24 g (1.26 mmol) of 5-bromo-2-thiophene-carboxaldehydeand 0.185 mL of triethylamine in 10 mL of acetonitrile was added with0.84 g (2.89 mmole) of 1,2-dimethylbenzothiazolium iodide. The reactionmixture was slowly heated to reflux for 16 hours. The initial color wasyellow. It turned red as temperature increased to 67° C. and it was darkgreen when the reaction was completed. The final product wasrecrystallized by dissolution in methanol and precipitation from ethylether; yield 70 mg: ¹ H NMR (300 MHz, DMSO-d₆) d 3.18, 3.37, 3.7, 4.2,4.3, 6.70-8.5 ppm; ¹³ C NMR (75.6 MHz, DMSO-d₆) d 18.1, 37.2, 117.9,125.4, 125.6, 129.1, 130.4 ppm. UV-vis (methanol) l_(max) (e, mol⁻¹ dm³cm⁻¹) 707 (14,000) nm; ESI-MS m/z for M⁺ is 419 which corresponds to C₂₇H₃₁ N₂ S₃.

Example II

Synthesis of Dimethylbenzothiazolium monocarbofuran iodide

A mixture of 0.22 g (1.26 mmol) of 5-bromo-2-furan carboxyaldehyde (C₃H₃ BrO₂) and 0.18 mL of triethylamine in 10 mL of acetonitrile is addedwith 0.84 g (2.89 mmole) of 1,2-dimethylbenzothiazolium iodide. Thereaction mixture is slowly heated to reflux for 16 hours. The initialcolor is yellow. It rams red as temperature increased to 67° C. and itis dark green when the reaction is completed. The final product ispurified by recrystallization or chromatography.

Example III

Synthesis of Dimethylbenzothiazolium monocarbo-1-methylpyrrole iodide

A mixture of 0.24 g (1.26 mmol) of 5-bromo-1-methyl-2-pyrrolecarboxaldehyde and 0.185 mL of triethylamine in 10 mL of acetonitrile isadded with 0.84 g (2.89 mmole) of 1,2-dimethylbenzothiazolium iodide.The reaction mixture is slowly heated to reflux for 16 hours. Theinitial color is yellow. It turns red as temperature increases to 67° C.and is dark green when the reaction is complete. The final product ispurified by recrystallization or chromatography.

Although the invention has been described with respect to specificmodifications, the details thereof are not to be construed aslimitations, for it will be apparent that various equivalents, changesand modifications may be resorted to without departing from the spiritand scope thereof, and it is understood that such equivalent embodimentsare to be included therein.

What is claimed is:
 1. A diagnostic composition comprising a compound ofthe formula: ##STR4## wherein R¹, R², and R⁵ may be the same ordifferent and are selected from the group consisting of hydrogen, C₁-C₁₀ alkyl, C₁ -C₁₀ alkoxyl, hydroxyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀alkoxyalkyl, C₁ -C₁₀ aryl, carboxyl, C₁ -C₁₀ carboxylalkyl, halogen,nitro, C₁ -C₁₀ alkoxycarbonyl, mercapto, C₁ -C₁₀ mercaptoalkyl, C₁ -C₁₀alkylthio, sulfonate, and --(CH₂)_(m) --N(R⁶)(R⁷) wherein R⁶ and R⁷ areindependently hydrogen or C₁ -C₁₀ alkyl, C₁ -C₁₀ acyl, and R⁶ and R⁷ arecapable of forming 5, 6, or 7 membered rings which may optionally besubstituted with --O--, --NR⁸, or --S--; R³ is selected from the groupconsisting of C₁ -C₁₀ alkyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ alkoxyalkyl,C₁ -C₁₀ aryl, C₁ -C₁₀ carboxylalkyl, C₁ -C₁₀ alkyl sulfonate, mercaptoalkyl and --(CH₂)_(m) N(R⁶)(R⁷); R⁴ is selected from the groupconsisting of C₁ -C₁₀ carboxylalkyl and --(CH₂)_(m) N(R⁶)(R⁷); W¹ and W²may be the same or different and are selected from the group consistingof --S--, --O--, --Se--, --Te--, --NR⁸ and C(R⁹)(R¹⁰); p is about 1-10;and m is about 0-10; R⁸ is hydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀ alkoxy, C₁-C₁₀ mercaptoalkyl, hydroxyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ alkoxyalkyl,C₁ -C₁₀ aryl, C₁ -C₁₀ carboxylalkyl, C₁ -C₁₀ alkoxycarbonyl, C₁ -C₁₀alkylthio, and --(CH₂)_(m) --N(R⁶)(R⁷); R⁹ and R¹⁰ are independentlyhydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀ alkoxyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀alkoxyalkyl, C₁ -C₁₀ carboxyalkyl, C₁ -C₁₀ alkoxycarbonyl, and--CH₂)_(m) --N(R⁶)(R⁷); and X is --O--, --S--, --SO--, --SO₂ --, --Se--,--Te--, --NR⁸, or --C(R⁹)(R¹⁰).
 2. The composition of claim 1 whereinR¹, R² and R⁵ are independently hydrogen, C₁ -C₁₀ alkyl, hydroxyl, C₁-C₁₀ alkoxyl, carboxyl, halogen, nitro, sulfonate, or --(CH₂)_(m)--N(R⁶)(R⁷) wherein m is about 0-10; R³ C₁ -C₁₀ alkyl, C₁ -C₁₀hydroxyalkyl, C₁ -C₁₀ carboxyalkyl, or --(CH₂)_(m) --N(R⁶)(R⁷); R⁴ is C₁-C₁₀ carboxyalkyl, or --(CH₂)_(m) --N(R⁶)(R⁷); R⁶ and R⁷ areindependently hydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀ acyl, and hydroxyalkyl; Xis --O--, --NR⁸ or --S--; W¹ and W² are independently --S-- or--C(R⁹)(R¹⁰) wherein R⁹ and R¹⁰ are independently hydrogen, C₁ -C₁₀alkyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ carboxyalkyl or --(CH₂)_(m)--N(R⁶)(R⁷); m is about 1-10; and R⁸ is hydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀hydroxyalkyl, C₁ -C₁₀ carboxyalkyl or --(CH₂)_(m) --N(R⁶)(R⁷).
 3. Thecomposition of claim 2 wherein R¹ is hydrogen; R² is hydrogen; R³ ismethyl; R⁴ is C₁ -C₁₀ carboxyalkyl; R⁵ is hydrogen; W¹ is --S--; W² is--S--; and X is --O--.
 4. The composition of claim 2 wherein R¹ ishydrogen; R² is hydrogen; R³ is methyl; R⁴ is C₁ -C₁₀ carboxyalkyl; R⁵is hydrogen; W¹ is --S--; W² is --S--; and X is --NH--.
 5. Thecomposition of claim 2 wherein R¹ is hydrogen; R² is hydrogen; R³ ismethyl; R⁴ is --(CH₂)_(m) --N(R⁶)(R⁷); W¹ is --S--; W² is --S--; and Xis --S--.
 6. The composition of claim 2 wherein R¹ is hydrogen; R² ishydrogen; R³ is methyl; R⁴ is --(CH₂)_(m) --N(R⁶)(R⁷); W¹ is --S--; W²is --S--; X is --O--; R⁶ is hydrogen; and R⁷ is hydrogen.
 7. Thecomposition of claim 2 wherein R¹ is hydrogen; R² is hydrogen; R³ ismethyl; R⁴ is --(CH₂)_(m) --N(R⁶)(R⁷); W¹ is --S--; W² is --S--; X is--S--; R⁶ is hydrogen; and R⁷ is hydrogen.
 8. The composition of claim 2wherein R¹ is hydrogen; R² is hydrogen; R³ is methyl; R⁴ is --(CH₂)_(m)--N(R⁶)(R⁷); W¹ is --S--; W² is --S--; X is --NH--; R⁶ is hydrogen; andR⁷ is hydrogen.
 9. A method of imaging a patient comprising theadministration of a diagnostically effective amount of a compound of theformula: ##STR5## wherein R¹, R², and R⁵ may be the same or differentand are selected from the group consisting of hydrogen, C₁ -C₁₀ alkyl,C₁ -C₁₀ alkoxyl, hydroxyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ alkoxyalkyl, C₁-C₁₀ aryl, carboxyl, C₁ -C₁₀ carboxylalkyl, halogen, nitro, C₁ -C₁₀alkoxycarbonyl, mercapto, C₁ -C₁₀ mercaptoalkyl, C₁ -C₁₀ alkylthio,sulfonate, and --(CH₂)_(m) --N(R⁶)(R⁷) wherein R⁶ and R⁷ areindependently hydrogen or C₁ -C₁₀ alkyl, C₁ -C₁₀ acyl, and R⁶ and R⁷ arecapable of forming 5, 6, or 7 membered rings which may optionally besubstituted with --O--, --NR⁸, or --S--; R³ is selected from the groupconsisting of C₁ -C₁₀ alkyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ alkoxyalkyl,C₁ -C₁₀ aryl, C₁ -C₁₀ carboxylalkyl, C₁ -C₁₀ alkyl sulfonate, mercaptoalkyl and --(CH₂)_(m) N(R⁶)(R⁷); R⁴ is selected from the groupconsisting of C₁ -C₁₀ carboxylalkyl and --(CH₂)_(m) N(R⁶)(R⁷); W¹ and W²may be the same or different and are selected from the group consistingof --S--, --O--, --Se--, --Te--, --NR⁸ and --C(R⁹)(R¹⁰); p is about1-10; and m is about 0-10. R⁸ is hydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀alkoxy, C₁ -C₁₀ mercaptoalkyl, hydroxyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀alkoxyalkyl, C₁ -C₁₀ aryl, C₁ -C₁₀ carboxylalkyl, C₁ -C₁₀alkoxycarbonyl, C₁ -C₁₀ alkylthio, and --(CH₂)_(m) --N(R⁶)(R⁷); R⁹ andR¹⁰ are independently hydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀ alkoxyl, C₁ -C₁₀hydroxyalkyl, C₁ -C₁₀ alkoxyalkyl, C₁ -C₁₀ carboxyalkyl, C₁ -C₁₀alkoxycarbonyl, and --(CH₂)_(m) --N(R⁶)(R⁷); X is --O--, --S--, --SO--,--SO₂ --, --Se--, --Te--, --NR⁸, or --C(R⁹)(R¹⁰), and obtaining animage.
 10. The method of claim 9 wherein R¹, R² and R⁵ are independentlyhydrogen, C₁ -C₁₀ alkyl, hydroxyl, C₁ -C₁₀ alkoxyl, carboxyl, halogen,nitro, sulfonate, or --(CH₂)_(m) --N(R⁶)(R⁷) wherein m is about 0-10; R³is C₁ -C₁₀ alkyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ carboxyalkyl or--(CH₂)_(m) --N(R⁶)(R⁷); R⁴ is C₁ -C₁₀ carboxyalkyl or --(CH₂)_(m)--N(R⁶)(R⁷); R⁶ and R⁷ are independently hydrogen, C₁ -C₁₀ alkyl, C₁-C₁₀ acyl, and hydroxyalkyl; X is --O--, --NR⁸ or --S--; W¹ and W² areindependently --S-- or --C(R⁹)(R¹⁰) wherein R⁹ and R¹⁰ are independentlyhydrogen, C₁ -C₁₀ alkyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ carboxyalkyl, or--(CH₂)_(m) --N(R⁶)(R⁷); m is about 1-10; and R⁸ is hydrogen, C₁ -C₁₀alkyl, C₁ -C₁₀ hydroxyalkyl, C₁ -C₁₀ carboxyalkyl, or --(CH₂)_(m)--N(R⁶)(R⁷).
 11. The composition of claim 10 wherein R¹ is hydrogen; R²is hydrogen; R³ is methyl; R⁴ is C₁ -C₁₀ carboxyalkyl; R⁵ is hydrogen;W¹ is --S--; W² is --S--; and X is --O--.
 12. The method of claim 11wherein R¹ is hydrogen; R² is hydrogen; R³ is methyl; R⁴ is C₁ -C₁₀carboxyalkyl; R⁵ is hydrogen; W¹ is --S--; W² is --S--; and X is --NH--.13. The method of claim 12 wherein R¹ is hydrogen; R² is hydrogen; R³ ismethyl; R⁴ is C₁ -C₁₀ carboxyalkyl; R⁵ is hydrogen; W¹ is --S--; W² is--S--; and X is --S--.
 14. The method of claim 13 wherein R¹ ishydrogen; R² is hydrogen; R³ is methyl; R⁴ is --(CH₂)_(m) --N(R⁶)(R⁷);W¹ is --S--; W² is --S--; X is --S--; R⁶ is hydrogen; and R⁷ ishydrogen.
 15. The method of claim 14 wherein R¹ is hydrogen; R² ishydrogen; R³ is methyl; R⁴ is --(CH₂)_(m) --N(R⁶)(R⁷); W¹ is --S--; W²is --S--; X is --O--; R⁶ is hydrogen; and R⁷ is hydrogen.
 16. The methodof claim 15 wherein R¹ is hydrogen; R² is hydrogen; R³ is methyl; R⁴ is--(CH₂)_(m) --N(R⁶)(R⁷); W¹ is --S--; W² is --S--; X is --NH--; R⁶ ishydrogen; and R⁷ is hydrogen.